Bottom Line:
CR-Leptin males showed higher hypothalamic long-form leptin receptor (ObRb) mRNA levels, compared to control and CR animals.In CR females, leptin treatment reverted the increased number of cells in ARC and cell density in ARC and PVN, and reduced hypothalamic SOCS-3 mRNA expression to levels similar to controls.In conclusion, leptin supplementation throughout lactation is able to revert, at least partly, most of the developmental effects on hypothalamic structure and function caused by moderate maternal caloric restriction during gestation, and hence making this metabolic malprogramming reversible to some extent.

ABSTRACTA poor prenatal environment brings about perturbations in leptin surge and hypothalamic circuitry that program impaired ability to regulate energy homeostasis in adulthood. Here, using a rat model of moderate maternal caloric restriction during gestation, we aimed to investigate whether leptin supplementation with physiological doses throughout lactation is able to ameliorate the adverse developmental malprogramming effects exerted in offspring hypothalamus structure and function. Three groups of male and female rats were studied: the offspring of ad libitum fed dams (controls), the offspring of 20% calorie restricted dams during the first part of pregnancy (CR), and CR rats supplemented with physiological doses of leptin throughout lactation (CR-Leptin). Animals were sacrificed on postnatal day 25. Morphometric and immunohistochemical studies on arcuate (ARC) and paraventicular (PVN) nucleus were performed and hypothalamic expression levels of selected genes were determined. In CR males, leptin treatment restored, at least in part, the number of immunoreactive neuropeptide Y (NPY(+)) cells in ARC, the total number of cells in PVN, hypothalamic NPY, cocaine- and amphetamine-regulated transcript (CART) and suppressor of cytokine signalling-3 (SOCS-3) mRNA levels, and plasma leptin levels, which were decreased in CR animals. CR-Leptin males showed higher hypothalamic long-form leptin receptor (ObRb) mRNA levels, compared to control and CR animals. In CR females, leptin treatment reverted the increased number of cells in ARC and cell density in ARC and PVN, and reduced hypothalamic SOCS-3 mRNA expression to levels similar to controls. Leptin treatment also reverted the increased relative area of NPY(+) fibers in the PVN occurring in CR animals. In conclusion, leptin supplementation throughout lactation is able to revert, at least partly, most of the developmental effects on hypothalamic structure and function caused by moderate maternal caloric restriction during gestation, and hence making this metabolic malprogramming reversible to some extent.

pone-0081906-g001: Offspring parameters.A. Energy intake, anthropometric measurements, circulating parameters and leptin mRNA and protein levels in stomach in the offspring of rats with free access to standard chow diet (control), the offspring of 20% calorie restricted dams during the first 12 days of pregnancy (CR), and CR rats daily supplemented with physiological doses of leptin throughout lactation (CR-Leptin). Cumulative energy intake (from postnatal day 21 to 25) was expressed in kcal, and also referred to body weight and expressed in kcal/g. Body weight was measured on postnatal days 1, 21 and 25. The other parameters were determined on day 25. Leptin mRNA levels in stomach were measured by qRT-PCR and expressed as a percentage of the value of control male rats. Leptin levels in stomach were quantified by ELISA and expressed in pg/g tissue. Data are mean ± S.E.M. For cumulative food intake, body weight at different days, body length and body fat content, n = 16-17; for mRNA analysis, n = 10-11; for circulating parameters, n = 6-8. Each group is made up of animals coming from at least six different litters. Statistics: in case of interaction within each sex, data not sharing a common letter (a and b) are significantly different (a≠b) (p < 0.05; LSD posthoc one-way ANOVA test). B. Evolution of body weight during lactation. The arrow indicates the starting point of significant effects of maternal caloric restriction during gestation on body weight in male and female offspring (CR≠Controls; CR-Leptin≠Controls; p < 0.05; LSD posthoc one-way ANOVA test).

Mentions:
Data related with energy intake, body weight, body fat and body length measurements, circulating parameters and gastric leptin in the offspring at birth and/or after weaning are summarized in Figures 1A and B. Maternal caloric restriction during gestation did not affect body weight at birth. However, during the suckling period, CR pups gained less weight and exhibited lower body weight that their controls from postnatal day 6 onwards (p < 0.05; LSD post hoc one-way ANOVA test). Cumulative energy intake after weaning, from day 21 to day 25, was lower in CR animals with respect to controls (p < 0.05; LSD post hoc one-way ANOVA test), although no differences were found when this was referred to body weight. Leptin treatment throughout lactation had no demonstrable effects on body weight of suckling pups, or in post-weaned animals at this juvenile age. However, post-weaned CR-Leptin females showed cumulative food intake slightly higher to that of CR animals, but not different to that of controls (p < 0.05; LSD post hoc one-way ANOVA test). At any rate, no differences were found when food intake was referred to body weight.

pone-0081906-g001: Offspring parameters.A. Energy intake, anthropometric measurements, circulating parameters and leptin mRNA and protein levels in stomach in the offspring of rats with free access to standard chow diet (control), the offspring of 20% calorie restricted dams during the first 12 days of pregnancy (CR), and CR rats daily supplemented with physiological doses of leptin throughout lactation (CR-Leptin). Cumulative energy intake (from postnatal day 21 to 25) was expressed in kcal, and also referred to body weight and expressed in kcal/g. Body weight was measured on postnatal days 1, 21 and 25. The other parameters were determined on day 25. Leptin mRNA levels in stomach were measured by qRT-PCR and expressed as a percentage of the value of control male rats. Leptin levels in stomach were quantified by ELISA and expressed in pg/g tissue. Data are mean ± S.E.M. For cumulative food intake, body weight at different days, body length and body fat content, n = 16-17; for mRNA analysis, n = 10-11; for circulating parameters, n = 6-8. Each group is made up of animals coming from at least six different litters. Statistics: in case of interaction within each sex, data not sharing a common letter (a and b) are significantly different (a≠b) (p < 0.05; LSD posthoc one-way ANOVA test). B. Evolution of body weight during lactation. The arrow indicates the starting point of significant effects of maternal caloric restriction during gestation on body weight in male and female offspring (CR≠Controls; CR-Leptin≠Controls; p < 0.05; LSD posthoc one-way ANOVA test).

Mentions:
Data related with energy intake, body weight, body fat and body length measurements, circulating parameters and gastric leptin in the offspring at birth and/or after weaning are summarized in Figures 1A and B. Maternal caloric restriction during gestation did not affect body weight at birth. However, during the suckling period, CR pups gained less weight and exhibited lower body weight that their controls from postnatal day 6 onwards (p < 0.05; LSD post hoc one-way ANOVA test). Cumulative energy intake after weaning, from day 21 to day 25, was lower in CR animals with respect to controls (p < 0.05; LSD post hoc one-way ANOVA test), although no differences were found when this was referred to body weight. Leptin treatment throughout lactation had no demonstrable effects on body weight of suckling pups, or in post-weaned animals at this juvenile age. However, post-weaned CR-Leptin females showed cumulative food intake slightly higher to that of CR animals, but not different to that of controls (p < 0.05; LSD post hoc one-way ANOVA test). At any rate, no differences were found when food intake was referred to body weight.

Bottom Line:
CR-Leptin males showed higher hypothalamic long-form leptin receptor (ObRb) mRNA levels, compared to control and CR animals.In CR females, leptin treatment reverted the increased number of cells in ARC and cell density in ARC and PVN, and reduced hypothalamic SOCS-3 mRNA expression to levels similar to controls.In conclusion, leptin supplementation throughout lactation is able to revert, at least partly, most of the developmental effects on hypothalamic structure and function caused by moderate maternal caloric restriction during gestation, and hence making this metabolic malprogramming reversible to some extent.

ABSTRACTA poor prenatal environment brings about perturbations in leptin surge and hypothalamic circuitry that program impaired ability to regulate energy homeostasis in adulthood. Here, using a rat model of moderate maternal caloric restriction during gestation, we aimed to investigate whether leptin supplementation with physiological doses throughout lactation is able to ameliorate the adverse developmental malprogramming effects exerted in offspring hypothalamus structure and function. Three groups of male and female rats were studied: the offspring of ad libitum fed dams (controls), the offspring of 20% calorie restricted dams during the first part of pregnancy (CR), and CR rats supplemented with physiological doses of leptin throughout lactation (CR-Leptin). Animals were sacrificed on postnatal day 25. Morphometric and immunohistochemical studies on arcuate (ARC) and paraventicular (PVN) nucleus were performed and hypothalamic expression levels of selected genes were determined. In CR males, leptin treatment restored, at least in part, the number of immunoreactive neuropeptide Y (NPY(+)) cells in ARC, the total number of cells in PVN, hypothalamic NPY, cocaine- and amphetamine-regulated transcript (CART) and suppressor of cytokine signalling-3 (SOCS-3) mRNA levels, and plasma leptin levels, which were decreased in CR animals. CR-Leptin males showed higher hypothalamic long-form leptin receptor (ObRb) mRNA levels, compared to control and CR animals. In CR females, leptin treatment reverted the increased number of cells in ARC and cell density in ARC and PVN, and reduced hypothalamic SOCS-3 mRNA expression to levels similar to controls. Leptin treatment also reverted the increased relative area of NPY(+) fibers in the PVN occurring in CR animals. In conclusion, leptin supplementation throughout lactation is able to revert, at least partly, most of the developmental effects on hypothalamic structure and function caused by moderate maternal caloric restriction during gestation, and hence making this metabolic malprogramming reversible to some extent.